This invention relates to an anti-stress agent and functional food having effects of preventing and mitigating mental and physical symptoms caused by stress.
In the modern society, people undergoes various kinds of stress caused by highly advanced and complicated scientific technology, or drastically changing social circumstances. Particularly, in the internationalized society, complex human relationships are formed, causing mental stress. It has been reported that a variety of symptoms are caused by mental stress.
It is recognized that mental stress has a great influence on circulatory system and immune system. However, the scientific concept and definition of stress have not yet been well established, so that means of evaluation of stress still have many problems, combined with methodological difficulties. However, in the recent years, studies of stress have been made from the medical point of view.
For example, it is reported that when one undergoes stress, angiotensin II increases, and intracorporeal sodium due to sodium reabsorbancy becomes excess, which causes rise in blood pressure (Osamu Mobara et al.: Taisha, 28, 2, 323, 1991). Based on such findings, studies have been made on the effect of enalapril and alacepril, which are angiotensin converting enzyme inhibitors and used as antihypertensive agents, on hypertension caused by stress (The American Journal of Cardiology; 68, 15, 1362(1991), Internal Medicine; 32, 9, 691(1993)). However, it is considered that suffering stress not only causes rise in blood pressure, but also influences various factors to cause stomach ulcer, ischemic heart diseases, cerebrovascular diseases, hyperlipemia, or the like. Therefore, though stress is regarded as one of the causes of hypertension, it is not believed that the anti-stress effect is achieved merely by suppressing the rise in blood pressure.
As an agent for preventing and mitigating mental and physical symptoms caused by stress, chemically synthesized medicaments such as a tranquilizer, an antianxiety agent, and sleeping pills are presently used. However, these medicaments have habituation and side effect problems, so that it is not preferable to use them daily for the purpose of preventing mental and physical symptoms caused by stress. Accordingly, an anti-stress agent that can be taken repeatedly and daily without any problems with safety, and that can mitigate and prevent mental and physical symptoms caused by stress are desired and are under development. For example, there are proposals such as an anti-stress agent containing as an effective ingredient L-theanine contained in tea leaves (Japanese Laid-open Patent Application No. 6-100442), an anti-stress composition containing imidazole compounds such as anserine, valenine, n-methylhistidine, or r-methylhistidine (Japanese Laid-open Patent Application No. 9-20660), and anti-stress food containing a composition of glutathione and antioxidant (Japanese Laid-open Patent Application No. 8-275752). There is also a report on stress reducing effect of fragrance (Fragrance Journal: 1991-11, p44-49). However, there has not been reported that a tripeptide has the effect of mitigating and preventing mental and physical symptoms caused by stress.
It is an object of the present invention to provide an anti-stress agent and functional food which can fulfill the social demand as mentioned above, which can be taken repeatedly and daily without any problems with safety, and which can mitigate and prevent mental and physical symptoms caused by stress.
According to the present invention, there is provided an anti-stress agent comprising as an effective ingredient a tripeptide and/or a salt thereof having angiotensin converting enzyme inhibitory activity.
According to the present invention, there is also provided the anti-stress agent wherein said tripeptide is Ile-Pro-Pro and/or Val-Pro-Pro.
According to the present invention, there is further provided use of the tripeptide and/or the salt thereof for the manufacture of an anti-stress agent.
According to the present invention, there is also provided food having an anti-stress effect and comprising said anti-stress agent.
According to the present invention, there is further provided use of the tripeptide and/or the salt thereof for the manufacture of food having an anti-stress effect.
According to the present invention, there is further provided a method for producing the food having anti-stress effect comprising fermenting a medium containing a peptide and/or a protein including a sequence Ile-Pro-Pro and/or Val-Pro-Pro with lactic acid bacteria under conditions for a tripeptide Ile-Pro-Pro and/or Val-Pro-Pro to be produced in a resulting fermented medium.
According to the present invention, there is also provided a method for reducing stress comprising orally administering an effective amount of a tripeptide and/or a salt thereof having angiotensin-converting enzyme inhibitory activity.
The anti-stress agent of the present invention contains, as an effective ingredient, a tripeptide and/or a salt thereof having angiotensin converting enzyme inhibitory activity. In the present invention, xe2x80x9canti-stressxe2x80x9d effect means an activity to cause approximation of the conditions of a subject to the conditions without stress, e.g., an activity to reduce systolic and diastolic blood pressures which have been increased due to stress, and an activity to inhibit or prevent lowering in immunological function caused by stress, such as lowering in spleen cell response.
The tripeptide may be preferably selected from the group consisting of Ile-Pro-Pro, Val-Pro-Pro (abbreviated hereinbelow as IPP and VPP, respectively) and mixtures thereof that have angiotensin converting enzyme inhibitory activity.
The salt of the tripeptide may include pharmacologically acceptable salts, such as inorganic salts such as hydrochloride, sulfate and phosphate, and organic salts such as citrate, maleate, fumarate, tartrate and lactate.
The tripeptide may be produced by, e.g., fermentation with microorganism, enzyme hydrolysis, or chemical synthesis.
The fermentation with microorganism can be performed by culturing lactic acid bacteria in a medium of a food material containing a peptide and/or a protein including an amino acid sequence corresponding to the tripeptide, such as sequences Ile-Pro-Pro and/or Val-Pro-Pro.
The medium is preferably a food material containing a peptide and/or a protein including an amino acid sequence corresponding to the tripeptides. The food material may be milk, milk casein, corn, corn protein, wheat, wheat protein, soybean, de-fat soybean or soybean protein. The medium may further contain other ingredients such as yeast extract, vitamins and minerals, if necessary.
As the lactic acid bacteria, lactic acid bacteria of the genus Lactobacillus may be employed. For example, Lactobacillus helveticus, Lactobacillus delbruekii subsp. bulgaricus, Lactobacillus acidophilus, Lactobacillus fermentum or Lactobacillus casei subsp. casei maybe employed. Specifically, Lactobacillus helveticus ATCC55796, Lactobacillus delbruekii subsp. bulgaricus ATCC11842, Lactobacillus acidophilus ATCC4356, Lactobacillus fermentum ATCC14931 or Lactobacillus casei subsp. casei ATCC393 may be employed.
The fermentation may be performed by heat-sterilizing the medium, cooling the medium to a desired culturing temperature, and then inoculating the medium with a pre-cultured lactic acid bacteria starter. The inoculation amount of the lactic acid bacteria starter is preferably 105 to 107 cells of the lactic acid bacteria per 1 g of the medium. The culture temperature may be 20 to 50xc2x0 C., and preferably 30 to 45xc2x0 C. The culturing time maybe 3 to 48 hours, and preferably 6 to 24 hours. The culturing may be terminated when number of cells of the lactic acid bacteria reaches 108 cells/g or more and acidity of the lactic acid reaches 1 or more. The resulting cultured medium usually contains 0.1 to 100 xcexcg/g of IPP and/or VPP, although it depends on the material and composition of the medium.
The cultured medium itself containing live lactic acid bacteria may be used as the anti-stress agent of the present invention. Alternatively, the cultured medium may be used after heat-sterilizing, e.g., up to 80xc2x0 C. Further, the cultured medium may be used after being powdered by freeze drying, spray drying or drum dryer drying.
The cultured medium may be concentrated to purify the tripeptide before use as the anti-stress agent of the present invention. The concentration and purification may be carried out by subjecting the cultured medium to centrifugation and taking supernatant of the centrifuged liquid. The supernatant may further be subjected to electrodialysis, ion exchange resin treatment, hollow fiber membrane dialysis, reverse osmosis treatment, hydrophobic column chromatography, or combinations thereof, to obtain a further concentrated and purified tripeptide.
The enzyme hydrolysis for producing the tripeptide may include treating a food material containing a peptide and/or a protein including an amino acid sequence corresponding to the tripeptides such as Ile-Pro-Pro and Val-Pro-Pro, successively with proteinase and carboxypeptidase. The proteinase may be proteinase derived from microorganisms, from plants, or from animals, and may be prepared by a publicly known method. The carboxypeptidase may be carboxypeptidase derived from microorganisms, from plants, or from animals, and may be prepared by a publicly known method.
The chemical synthesis for producing the tripeptide may include publicly known organic synthesis. For example, the tripeptide may be obtained by preparing amino acids that are components of the objective tripeptide, protecting the amino group of each amino acid with a fluorenylmethoxycarbonyl group, successively reacting the amino acids protected with the fluorenylmethoxycarbonyl group in accordance with the amino acid sequence of the objective tripeptide by any conventional method, to obtain a tripeptide bound to a resin, and then removing the resin by any conventional method and purifying the tripeptide.
The content of the tripeptide in the anti-stress agent of the present invention is not particularly limited as long as it enables administration of the effective amount of the tripeptide to be discussed later. The content of the tripeptide may usually be 0.001 to 1 wt % of the agent.
The anti-stress agent of the present invention may contain, in addition to the tripeptide and/or the salt thereof, other additives such as sugars, proteins, lipids, vitamins, minerals, flavoring agents and coloring agents.
The anti-stress agent of the present invention may be administered to humans or animals. The administration may be performed orally or intravenously, with the oral administration being preferable.
Effective administration amount of the anti-stress agent of the present invention may usually be, for example, in a range of 0.1 to 40 mg/kg body weightxc2x7day in terms of the tripeptide for oral administration, in order to achieve the effect of the present invention such as mitigation and prevention of stress in human. However, the anti-stress agent may be administered in an amount exceeding this range.
When the anti-stress agent of the present invention is administered to a subject, changes in various physiological indices in response to stress, such as rise in blood pressure, lowering in immunological functions, becomes smaller than that in a case in which the present agent is not administered, and approximates to that in a case in which stress is not loaded.
The functional food of the present invention contains the anti-stress agent. Taking this functional food prevents or mitigates the stress.
The content of the anti-stress agent in the functional food of the present invention is not particularly limited as long as it is in a range to enable the tripeptide and/or the salt thereof in the functional food achieves the anti-stress effect. The functional food may usually contain the anti-stress agent in an amount to make the content of the tripeptide and/or the salt thereof in the functional food be 0.001 to 0.1 wt %.
By taking the functional food of the present invention in an amount ranging from 0.1 to 40 mg in terms of the tripeptide per kg body weightxc2x7day, preferable effects of the present invention may be obtained. However, the functional food may be taken in an amount exceeding this range.
The functional food of the present invention may include yogurt, milk-containing acidified beverages, cheese, processed foods and beverages containing fermented sour milk or health foods, and may be in the form of solid such as powders, granules and tablets, or of fluid such as paste, gel and liquid.
The tripeptide, the effective ingredient of the anti-stress agent and the functional food of the present invention may be obtained by fermenting a food material with lactic acid bacteria, and thus is quite safe.
Since the anti-stress agent and the functional food of the present invention contain the tripeptide, they are quite safe, and repeated and daily intake thereof can mitigate and prevent mental and physical symptoms caused by various kinds of stress.